dsRNA阻断大鼠骨髓源性神经干细胞Hes5表达的实验研究  

Blocking with double-stranded RNA of the expression of Hes5in rat bone marrow-derived neural stem cells

作　　者：王海燕[1] 徐如祥[1] 姜晓丹[1] 罗深秋[2] 

机构地区：[1]第一军医大学珠江医院全军神经医学研究所,广东广州510282 [2]第一军医大学细胞生物学教研室学,广东广州510515

出　　处：《第一军医大学学报》2004年第1期35-38,41,共5页Journal of First Military Medical University

基　　金：国家自然科学基金(3027049);军队及广东省科技厅"十五重大项目(01z054)~~

摘　　要：目的 观察外源性短双链RNA(dsRNA)在转录后水平即mRNA水平降低基因表达的效率，并对其影响因素进行初步探讨。方法 将体外分离培养的大鼠骨髓基质细胞，通过由本实验室配制的特殊培养基诱导成神经干细胞，应用人工合成的dsRNA于不同浓度转染神经干细胞，Western bloting检测dsRNA是否能阻断转录因子Hes5的表达，0．5％锥虫蓝法测定各浓度组中培养细胞活力。结果 200、300nmol／L浓度的dsRNA能特异有效地阻断Hes5的表达，而50～200nmol／L浓度的dsRNA有利于干细胞存活。结论 dsRNA能在神经干细胞内启动RNA干扰作用，适宜浓度的dsRNA能在特异有效地阻断内源性基因的表达的同时，最大限度地保持细胞活力。Objective To examine the efficiency of exogenous small d ouble-stranded RNA (dsRNA) in knocking down the gene expression at the post-tra nscription level, and investigate the factors that may influence the transfectio n. Method The bone marrow stromal cells of SD rat were separated and cultured i n vitro, followed by induction of the cells to evolve into neural stem cells us ing special culture medium prepared by our laboratory. Synthetic dsRNA was then transferred into the cells at varied concentrations, and the results were analy zed by Western blotting. Results The concentrations ranging from 200 to 300 nmol /L were optimal for specifically blocking the expression of Hes5, whereas the su itable concentrations for the cell survival were between 50 and 200 nmol/L. Con clusion dsRNA is capable of triggering RNA interference in neural stem cells, a nd at appropriate concentration, it may specifically and effectively knock down endogenous gene expression without sacrificing the viability of the cells.

关 键 词：DSRNA 大鼠 骨髓源性神经干细胞 Hes5 实验 外源性短双链RNA 基因表达 

分 类 号：R33[医药卫生—人体生理学]